This disclosure relates to methods of determining the antibiotic susceptibility of microbial cells in a sample. This disclosure also relates to methods of measuring RNA in microbial cells.
Emergence of drug resistant pathogens is a global healthcare crisis that is forcing physicians to treat common infectious diseases with ever more expensive, potent, and sometimes more toxic antibiotics. Unfortunately, pharmaceutical development of new antibiotics has rapidly declined, resulting in a lack of new agents to treat some organisms that are multi-drug resistant. Mainstream clinical microbiology is slow and expensive because it still relies on bacterial growth for colony formation on agar plates, a time-consuming, labour-intensive method requiring skilled technicians who are increasingly in short supply. Antibiotic susceptibility data are typically not available for 2-3 days after specimen acquisition, which is too late to meaningfully impact antibiotic selection. New rapid clinical microbiology methods are urgently needed that can perform identification and antibiotic susceptibility testing (AST) directly on pathogens found in clinical specimens, providing clinicians with real-time information to manage infectious diseases.
In the absence of an expeditious microbiologic diagnosis, clinicians typically initiate “empiric” antibiotic treatment, meaning that antibiotics are chosen based on knowledge of potential organisms and their antibiotic resistance patterns. Empiric antibiotics for bacteremia are typically broad-spectrum antibiotics to treat a wide variety of possible bacterial pathogens. Overuse of broad-spectrum antibiotics contributes to the emergence of antibiotic resistance by applying selective pressure to the patient's microbiota and favoring colonization by resistant organisms. For example, the common use of vancomycin and piperacillin-tazobactam as empiric therapy has contributed directly to the widespread emergence of vancomycin-resistant enterococci (VRE) and extended-spectrum beta-lactamase (ESBL) producing E. coli and Klebsiella pneumoniae organisms. In contrast, fungemia is frequently not suspected or treated until patients receiving antibacterial agents do not respond or experience clinical deterioration. Evidence suggests that the use of early, effective antibiotic therapy improves patient outcomes and shortens hospital length of stay. Therefore, the ability to quickly identify the causative organism(s) and administer appropriate antibiotic therapy should result in improved patient outcomes as well as reduce overall costs to the healthcare system.